Microwave-assisted chromatography preparation

ABSTRACT

An instrument and associated method are disclosed for preparing samples for column chromatography. The method includes the steps of applying microwave energy to a sample composition containing at least one solvent to encourage a chemical reaction and generate desired products, thereafter mixing an absorbent media with the sample to absorb the solvent, the media being compatible with liquid chromatography that will separate the expected products, being chemically inert to the expected products, and being added in an amount sufficient to provide a substantially dry mixture of the media and the sample, but less than an amount that overly broadens the resolution of the sample during liquid chromatography, thereafter applying microwave energy to the dry mixture of the media and the sample to thereby encourage the solvent to evaporate under the influence of the microwave energy, and thereafter adding the dry mixture of the media and the remaining sample to a liquid chromatography column and separating the components of the remaining sample for identification and purification purposes.

RELATED APPLICATIONS

This is a divisional application of Ser. No. 10/923,243 filed Aug. 20,2004 and now U.S. Pat. No.______.

FIELD OF THE INVENTION

The present invention relates to microwave-assisted flashchromatography, and in particular relates to a microwave instrument thatoffers particular advantages useful for sample preparation andpurification via chromatography.

BACKGROUND OF THE INVENTION

The present invention relates to devices for microwave-assistedchromatography. As generally recognized in the chemical arts, manychemical reactions can be initiated or accelerated by increasing thetemperature—i.e. heating—the reactants. Accordingly, carrying outchemical reactions at elevated (i.e., above ambient) temperatures is anormal part of many chemical processes.

The benefit of using controlled microwave energy for elevating thetemperature of a chemical reaction is well known. For example, U.S. Pat.No. 6,753,517 to Jennings, incorporated entirely herein by reference,discloses a microwave-assisted chemical synthesis instrument usingtightly controlled microwave energy.

More recently, researchers have applied microwave assisted chemistry tothe technique of chromatography. Chromatography in the present contextincludes liquid and gas chromatography. Of the two, the instrument ofthe present invention primarily relates to liquid chromatography,particularly as it pertains to solvent evaporation and samplepreparation for elution.

Liquid chromatography is a technique utilized in both preparative andanalytical chemistry. Liquid chromatography comprises a stationary phaseinteracting with a mobile phase. Typically, the stationary phase is asurface-active powder such as silica, alumina, or an inertsize-separating material like a gel-permeation chromatography packing,or the like. This powder is contained in a chromatographic column. Inpreparative chemistry, the mobile phase generally consists of a reactionsolvent and a chemical to be identified, analyzed, or purified. This iscollectively referred to as a sample. The mobile phase carrying thesample is caused to migrate through the stationary phase. Depending onhow the sample interacts with the surface characteristics of thestationary phase, different compounds will migrate through thestationary phase at different rates. Preparative chemistry is useful foridentifying and purifying various chemical components in the sampleusing analytical chemistry. Analytical chemistry utilizes carriersolvents to move the sample through the stationary phase.

In some instances, the reaction solvent is not ideal for the furtherpurification of the compound dissolved therein. Residual reactionsolvent in the stationary phase can result in the problem of poorseparation and recovery of the desired components during elution.

One solution to this problem is to utilize a rotary evaporator. Thisprocedure, however, is time consuming and not entirely effective. Forexample, the use of larger chromatography columns having more stationaryphase will require one or more hours to evaporate the reaction solvent.

Another solution to this problem is to apply heat with or withoutvacuum. This procedure, however, may result in the degradation of thestationary phase or melting of the chromatography column material.

Another solution to the problem of residual reaction solvent in thestationary phase is to flash away the solvent using microwave energy.Microwave energy is used to vaporize liquid in U.S. Pat. No. 4,330,946to Courneya. The Courneya '946 patent discloses microwave energy tovaporize liquid from agricultural material, such as grain. The Coumeya'946 patent further discloses the use of air inlets for introducing airto sweep across the material, aerosolize the vapor and carry it away viaa vacuum pump. The Coumeya '946 patent also states the need for anauger-driven process to insure proper agitation of the material, as wellas to move the material through the apparatus. A heat reclaim mechanismassists the microwave energy drying process.

As previously stated, microwave energy has been applied to the techniqueof chromatography. For example, U.S. Pat. No. 6,029,498 to Walters etal. discloses the incorporation of microwave absorbing material into thechromatography column itself or positions adjacent the column. Thechromatography sample contained therein is heated by the microwaveabsorbing material via conduction or convection. The Walters '498 patentfurther discloses, however, the use of a heated chromatography columnfor the purpose of enhancing the speed of separation and the consistencyof elution times.

Another example of the application of microwave energy to liquidchromatography is demonstrated in U.S. Pat. No. 6,630,354 to Stone. TheStone '354 patent discloses a method of using microwave-induceddielectric polarization to enhance the diffusivity of a liquid or asupercritical fluid mobile phase in chromatography, while havingessentially no effect on other physical properties of the mobile phase.The primary focus of the Stone '354 patent, however, is to increase thediffusivity of the reaction solvent and to combine the advantages ofliquid and gas chromatography. Heating the reaction solvent to increaseits diffusivity could be hazardous if the solvent is combustible. Toovercome this, the Stone '354 patent discloses the use of microwavepulses. The Stone '354 patent further states that a microwave apparatuscapable of delivering very short pulses of radiation is not available,and instead teaches the use of a conventional, non-cycling microwaveoven.

Yet another example of the application of microwave energy to liquidchromatography is demonstrated in U.S. Pat. No. 6,649,051 to Jamalabadiet al. The Jamalabadi '051 patent discloses a method of processing asample into a flow-through device containing a porous solid media andthereafter subjecting the device to microwave energy. As in the Stone'354 patent, the Jamalabadi '051 patent discloses the use of aconventional microwave oven and further discloses the use of a moreprecise microwave power source, such as one manufactured by CEMCorporation of Matthews, N.C., USA, the assignee of the presentinvention.

Yet another example of the application of microwave energy to liquidchromatography is demonstrated in U.S. Pat. Application Publication No.20030205456 to Jamalabadi et al. The Jamalabadi application discloses amethod of processing a sample comprising introducing a sample in aflow-through device containing a porous solid media therein. Theflow-through device is defined as having walls and having an inlet endand an outlet end; a solid porous media disposed within the flow-throughdevice including attached active components. After introducing a sampleinto the flow-through device, the device is subjected to a radiatedenergy source, such as microwave energy, prior to further chromatographysteps.

Furthermore, none of the above-referenced patents or applicationsteaches or suggests the use of a dry-loading technique. A typicaldry-loading technique includes addition of a given amount ofchromatography media, for example silica or alumina, to a vessel holdingthe sample. The sample is preferably dissolved in a reaction solvent.Following absorption of the sample by the chromatography media, thereaction solvent is evaporated and evacuated, leaving driedchromatography media containing the sample. The dried chromatographymedia is then dry-loaded, either manually or automatically, onto aseparation chromatography column for separation and identification ofthe sample components.

The advantage of a dry-loading technique lies in the flexibility itallows for evaporation of the reaction solvent. Accordingly, dry-loadingstreamlines the subsequent separation and analysis of the samplecomponents. The dry-loading technique is infinitely adjustable for agiven sample volume, as opposed to the limitations imposed by preloadinga specific amount of chromatography media into a flow-through device, ora sample module, or the like. For example, if the flow-through devicecontains too little chromatography media for a given amount of reactionsolvent, the chromatography media will not sufficiently absorb thesolvent. Subsequent heating of the chromatography media to evaporate thesolvent will result in a highly viscous mixture from which the samplecannot be salvaged. If the flow-through device contains too muchchromatography media for a given amount of reaction solvent, subsequentseparation and identification chromatography efforts will be confoundedby the excess media, resulting in wide, overlapping peaks having poorresolution.

OBJECT AND SUMMARY OF THE INVENTION

Therefore, it is an object of the invention to provide a microwaveinstrument suitable for microwave assisted chemistry, includingaccelerated chemical synthesis and sample preparation for flashchromatography and liquid chromatography. The invention meets thisobject with an instrument for carrying out a reaction in reactionsolvents in a microwave transparent reaction vessel followed byevaporating and evacuating reaction solvents from the chromatographymedia. The invention further includes a source of microwave radiationand a microwave cavity in wave communication with the source forapplying microwave energy to a sample. The sample includes a solid phasechromatography media and a solvent. The invention further includes avessel for holding the sample in the microwave cavity for evaporatingthe solvent when microwaves are applied to the vessel, and a gas pump ingas communication with the vessel for moving gases through the vesselduring the application of microwaves to the vessel to facilitate theevaporation of the reaction solvent from the chromatography media.

It is further an object of the present invention to provide a microwaveassisted chromatography sample preparation instrument, including amicrowave source, and a mechanism for controlling the application ofmicrowave energy from the microwave source. The instrument furtherincludes a vessel in wave communication with the microwave source forevaporating solvent, a gas inlet tube and a gas exit tube about thevessel for allowing the influx of make-up gas and the evacuation ofsolvent, respectively, and a gas pump in physical communication with thevessel to facilitate the evaporation of solvent. The gas pump, gas inlettube, and gas exit tube create a gas flow through the evaporation vesselto evaporate the solvent from the chromatography sample during andbetween applications of microwave energy.

It is yet another object of the present invention to provide a method ofpreparing samples for column chromatography, especially flashchromatography, comprising adding a sample including reaction solvent toa solid phase chromatography medium and applying microwave energy to thesample while concurrently providing a gas flow over and around thesample. The solvent is thereby encouraged to evaporate under theinfluence of the microwave energy and the flowing gas.

It is yet another object of the present invention to provide a methodfor microwave assisted chromatography sample preparation, includingmixing a sample dissolved in a reaction solvent with a solid phasechromatography medium, applying microwave energy to the sample toevaporate the solvents therein, and create a gas flow through a vesselto evacuate the vaporized solvents. The invention further provides formonitoring the temperature of the chromatography media using temperaturesensors and moderating the temperature of the sample. A microprocessoris used to control the steps of applying microwave energy, moderatingthe temperature of the sample, and creating the gas flow. The step ofcreating a gas flow further includes creating a vacuum in the vessel.

It is yet another object of the present invention to provide a method ofpreparing samples for column chromatography, such as flashchromatography, including applying microwave energy to a samplecomposition containing at least one solvent to encourage a chemicalreaction and generate desired products. Thereafter, the method includesmixing an absorbent media with the sample to absorb the solvent. Themedia is compatible with liquid chromatography that will separate theexpected products, is chemically inert to the expected products, and isadded in an amount sufficient to provide a substantially dry mixture ofthe media and the sample, but less than an amount that overly broadensthe resolution of the sample during liquid chromatography. The methodthereafter provides for applying microwave energy to the dry mixture ofthe media and the sample to thereby encourage the solvent to evaporateunder the influence of the microwave energy. The method of the inventionthen includes adding the dry mixture of the media and the remainingsample to a liquid chromatography column and separating the componentsof the remaining sample for identification and purification purposes.

It is yet another object of the present invention to provide a method ofpreparing samples for column chromatography as described, including theaddition of scavenging compositions, catalysts, and coupling reagents.

It is yet another object of the present invention to provide a method ofdry-loading samples for column chromatography.

It is further an object of the invention to provide the previouslydiscussed aspects while monitoring the temperature of the sample.

It is further an object of the invention to provide the previouslydiscussed aspects under the electronic control of a microprocessor, andadditionally providing that the microprocessor moderate the applicationof microwave energy based upon the monitored temperature.

The foregoing and other objects and advantages of the invention and themanner in which the same are accomplished will become clearer based onthe followed detailed description taken in conjunction with theaccompanying drawings in which:

DESCRIPTION OF THE DRAWINGS

FIG. 1 is a perspective view of the invention showing the instrumentwith the vessel placed in the microwave cavity.

FIG. 2 is a perspective view of the vessel and its components.

FIG. 3 is a cutaway diagram of the invention showing the instrument withthe vessel placed in the microwave cavity.

FIG. 4 is a perspective view of the invention with respect to a methodembodiment of the invention.

FIG. 5 is a further perspective view of the invention with respect to amethod embodiment of the invention.

FIG. 6 is a perspective view of the invention showing the back of theinstrument.

DETAILED DESCRIPTION OF THE INVENTION

In a first embodiment, the invention is a sample preparation instrumentfor flash chromatography. The instrument 10 is broadly illustrated inFIG. 1. The instrument 10 includes a microwave instrument housing 11typically made of rugged plastic. The housing 11 protects internalcomponents described herein. The housing 11 is vented with slottedapertures 14 to facilitate cooling of the internal components. Theinstrument 10 includes a microwave cavity 19 and a vessel 20 for holdingreactants, reaction solvent, products, and absorbent chromatographymedia. Further illustrated in FIG. 1 are communication devices for theinstrument 10, specifically a keyboard 16 and display 17. The instrument10 further includes at least one auxiliary port 12 for additionalattachments, such as a pressure attachment or an evaporation attachment.Additional parts of the vessel 20 shown in FIG. 1 include an adaptor 26,a gas inlet tube 28, a gas outlet tube 29, and a fitting 33 to connectthe tubes to peripheral devices explained herein.

FIG. 2 depicts the vessel 20 for holding reactants, reaction solvent,products, and absorbent chromatography media (i.e., compositions). Theillustrated vessel 20 is a test tube shaped device having a wall 21formed of a microwave transparent material defining an interior chamber22 to hold the composition. The vessel is not, however, limited tospecific vessel shapes and other vessel shapes (e.g., round-bottomflasks) can be incorporated as desired or necessary. The vessel 20includes a top portion 25. The top portion 25 further includes anadaptor 26 having an O-ring 27 for connecting and sealing the adaptor 26to the vessel wall 21. A hollow chemical resistant gas inlet tube 28projects into the vessel 20, allowing gas to flow into the chamber 22.Similarly, a hollow chemical resistant vent tube 29 allows gas,particularly solvent vapor, to exit the chamber 22. In this manner, thegas inlet tube 28 and the gas outlet tube 29 allow make-up gas into thevessel 20 while allowing solvent to evacuate the vessel 20. The topportion 25 further includes a frit (not shown) for containing thecomposition within the vessel chamber 22 during gas flow. The frit ismade of a chemical resistant media sufficiently porous to allow the flowof gas. The frit is disc-shaped having an opening to allow the gas inlettube 28 to pass therethrough.

In preferred embodiments the interior chamber 22 has a volume of atleast about 2.5 milliliters, which is a convenient size for bench-topexperiments. It will be understood that the invention is notvolume-limited, and that larger vessels can be used as may be necessaryor desired. Suitable microwave transparent materials are well known tothose of ordinary skill in the art, and include, for example, quartz,glass, and PYREX®.

The inlet tube 28 and the outlet tube 29 may have a fitting 33,generally denoted herein, for connecting the tube to another piece ofequipment such as a vacuum pump (see FIG. 3) or gas reservoir.Appropriate fittings 33 are known to those of ordinary skill in the artand include, for example, threaded fittings, valves, quick-connectfittings, and hose clamps.

FIGS. 1 and 3 show the microwave cavity 19 holding the vessel 20. In apreferred embodiment, the cavity 19 surrounds at least a portion of thevessel chamber 22. With respect to FIG. 3, the instrument's 10 variousinternal components are illustrated. FIG. 3 shows a microwave source 40,a waveguide 41, a stir motor 42, a fan 43 along with the fan housing 44,and various electronics.

A microwave source 40, as will be known to those of ordinary skill inthe art, can be microwave generating devices such as magnetrons,klystrons, and solid state devices. Microwaves travel from the source 40through the waveguide (schematically illustrated at 41) to the microwavecavity 19. The composition in the reaction vessel 20 absorbs themicrowave energy as it enters the cavity 19. In this manner, thereaction cavity 19 is in microwave communication with the microwavesource 40. In a preferred embodiment, the source 40 applies microwaveenergy to the reactants and the reaction solvent in the vessel 20 toaccelerate the conversion of reactants to products.

Following product formation, an absorbent media (not shown) is mixedwith the sample, typically by adding the media to the vessel 20 toabsorb the reaction solvent and products. The absorbent media ispreferably a chromatography media selected from the group consisting ofsilica, sand, alumina, and diatomaceous earth. Microwave energy isfurther applied to evaporate the reaction solvent. During theapplication of microwaves, a gas pump (schematically designated at 46)in gas communication with the vessel 20, preferably in communicationwith the gas exit tube 29, moves gases through the vessel 20 therebyfacilitating the evaporation of reaction solvent from the chromatographymedia. In preferred embodiments, the gas pump 46 is a vacuum pump forpulling gases from the vessel and thereby reducing the gas pressure overthe sample. Although a “hard” vacuum (e.g., approaching 0 Torr) is notrequired, the vacuum pump is a preferred device for reducing thepressure by the desired or necessary amount. In a preferred embodimentshown in FIG. 2, the gas inlet tube 28 includes a chemical resistanttube projecting into the vessel chamber 22 to carry make-up air near thebottom of the chromatography media inside to ensure thorough drying ofthe media. In this manner, the gas pump 46 maintains a vacuum in thevessel 20, specifically the vessel chamber 22, to facilitate theevaporation of the reaction solvent.

Alternatively, the gas pump 46 may be in physical communication with thegas inlet tube 28. In this arrangement, the pump 46 maintains pressurein the vessel 20, specifically the vessel chamber 22, to facilitate theevaporation of the reaction solvent from the chromatography media. Inthis embodiment, an inert gas is used to maintain the pressure in thechamber 22. Inert gas in this context refers to a gas that will notadversely react under the conditions in the vessel chamber 22. Forexample, a gas is chosen that will not ignite or otherwise combust withthe composition in any given reaction state within the chamber 22. Inmany cases, inert gas can be selected from the group consisting of air,nitrogen, and argon. Alternatively, a pressurized gas reservoir (notshown) can replace the pump 46. In such cases a gas regulator (notshown) between the gas reservoir and the vessel 20 controls the flow ofmake-up gas.

The waveguide 41 is constructed of a material that reflects microwavesinward and prevents them from escaping in any undesired manner.Typically, such material is an appropriate metal which, other than itsfunction for confining microwaves, can be selected on the basis of itscost, strength, formability, corrosion resistance, or any other desiredor appropriate criteria. In preferred embodiments of the invention, themetal portions of the waveguide 41 and cavity are formed of stainlesssteel.

As is the case with other kinds of chemistry, it is advantageous inmicrowave assisted organic chemistry to stir and mix the composition inthe vessel chamber 22. This is accomplished, for example, using a motor42 to drive a magnetic stirrer, such as described in U.S. PatentApplication Publication No. 20030170149, the contents of which arehereby entirely incorporated by reference.

The fan 43 serves to cool the electronics and the microwave source 40portions of the instrument 10, as well as helping to keep the reactioncavity 39 from becoming overheated in the presence of ongoing chemicalreactions. Other than having the capacity to appropriately cool theinstrument and the cavity, the nature or selection of the fan 43 can beleft to the individual discretion of those with skill in this art. In atypical embodiment, the fan 43 is mounted in a housing 44 to direct theflow of air across the electronics and the microwave source 40 to coolthem more efficiently.

In a preferred embodiment, the instrument 10 includes a temperaturesensor 35 and a pressure transducer 37 for measuring the temperature ofthe sample (i.e., composition) and for sensing the pressure inside thevessel 20, respectively. An appropriate temperature sensor can beselected from the group consisting of infra red detectors, ultravioletdetectors, and fiber optic sensors. In addition, the vessel top portion25 is designed to withstand the increased pressure generated by somemicrowave assisted organic reactions. The vessel top portion 25 mayinclude a pressure-resistant closure such as described in previouslyincorporated Publication No. 20030170149. In this embodiment, the vesseltop portion 25 may be held in place on the cavity 19 with a retainingring 38.

The instrument 10 includes a mechanism for adding chromatography mediato the vessel 20 to absorb the reaction solvent and the products. Themechanism permits manually adding about an equal volume ofchromatography media. Alternatively, the mechanism includesautomatically adding about an equal volume of chromatography media.

FIG. 3 also shows the microprocessor 48. The electronics carried by themicroprocessor 48 are generally well understood in their nature andoperation. With respect to the present instrument, the microprocessorfirst controls the power from a given source, usually a wall outletcarrying standard current. The microprocessor also controls theoperation of the device in terms of turning the microwave source 40 onor off, and in processing information received from the ongoing chemicalreaction, in particular pressure and temperature. In turn, the processoris used to control the application of microwaves, including startingthem, stopping them, or moderating them, in response to the pressure andtemperature information received from the sensors previously described.The use of processors and related electronic circuits to controlinstruments based on selected measured parameters (e.g., temperature andpressure) is generally well understood in this and related arts.Exemplary (but not limiting) discussions include Dorf, The ElectricalEngineering Handbook, Second Ed., (1997) CRC Press LLC.

With respect to the instrument and method embodiments of the invention,the automated aspects of the instrument 10 are controlled by themicroprocessor 48. This includes the control of simple peripheraldevices such as the gas pump 46 and more complex devices including, butnot limited to, CEM Corporation's EXPLORER™ and NAVIGATOR™ machines. SeeFIGS. 4 and 5. In a preferred embodiment, the invention is a modulardevice adapted to control peripheral devices while utilizing microwaveenergy to accelerate the process. Therefore, the preferred embodiment ofthe invention is simultaneous microprocessor control of the instrument10, including the application of microwave energy, moderating theapplication of microwave energy based on the monitored temperature orpressure from the previously mentioned sensors, peripheral devices inelectronic and physical communication with the instrument 10, and therespective methods incorporated by various combinations of theinstrument 10 and peripheral devices.

In another aspect, the invention is a microwave assisted chromatographysample preparation instrument 10. Referring to FIG. 3, the instrument 10includes a microwave source 40, a mechanism for controlling theapplication of microwave energy from the source 40, and a vessel 20 inwave communication with the source 40 for evaporating solvent.

The mechanism for controlling the application of microwave energy is themicroprocessor 48. In one embodiment of the invention, controlling theapplication of microwave energy applies the energy in pulses to avoidoverheating the sample in the vessel 20. An exemplary method formoderating the application of microwave energy is to incorporate avariable power supply such as disclosed in commonly assigned U.S. Pat.No. 6,084,226. Pulsing the microwave energy helps control the by-productof thermal heat. This decreases the likelihood of damaging or destroyingthe absorbent chromatography media and the reaction products absorbedtherein.

The instrument is also useful if a vessel other than that depicted inFIG. 2 is used. For example, U.S. Pat. Application Publication No.20030205456 to Jamalabadi et al., discloses a preloaded chromatographymodule (i.e., sample collector) having a plastic casing. The use of areaction solvent with a high boiling point, such as dimethylsulfoxide(DMSO) or dimethylformamide (DMF), will require a large amount ofmicrowave energy to evaporate the solvent. The heat generated duringevaporation could melt the plastic casing and ruin the sample. Thevariable power supply mechanism discussed above solves this problem bycontrolling the input of energy while monitoring the temperature of thesample.

With respect to the previously described elements of the invention, theinstrument 10 further includes a gas inlet tube 28 about the vessel 20for allowing make-up gas into the vessel 20, a gas exit tube 29 aboutthe vessel 20 for allowing solvent to evacuate the vessel 20, and a gaspump 46 in physical communication with the vessel 20 to facilitate theevaporation of solvent. In this embodiment, the gas pump 46, the gasinlet tube 28, and the gas exit tube create a gas flow through thevessel 20 to evaporate the solvent from the chromatography sample duringand between applications of microwave energy.

The instrument 10 includes the advantage of the previously discussedintegration of microprocessor control over other electronics andperipheral devices.

In another aspect, the invention is a method of preparing samples forcolumn chromatography. The method includes the step of mixing a sample,including a reaction solvent, with a solid phase chromatography mediumand applying microwave energy to the sample while concurrently providinga gas flow over and around the sample. The solvent is thereby encouragedto evaporate under the influence of the microwave energy and the flowinggas.

As previously discussed, the preferred embodiment of the method of theinvention (and necessarily the instrument and peripherals that carry outthe method) includes microprocessor control.

An example is shown in FIG. 4. One embodiment of the method of theinvention is carried out using the instrument 10 previously describedwith the EXPLORER™peripheral attachment 55. The sample, includingreaction solvent, is mixed with the chromatography medium in the samplecollector 56. In this embodiment, the sample collector 56 is a singleuse chromatography module, typically a plastic cylindrical casepre-loaded with chromatography media. In this arrangement, the sample isadded to the media already in the collector 56. Although different fromthe multi-use vessel 20 previously described, the method of the instantinvention is adaptable for use of the sample collector 56 in appropriatesample holder racks 57. The EXPLORER™ attachment 55 lowers the samplecollector 56 into the cavity 19 where the sample collector 56 is exposedto microwave energy. The instrument 10, in conjunction with a gas pump(not shown) concurrently provides a gas flow over and around the sampleto evaporate the solvent as described.

The chromatography preparation method utilizes microprocessor controlover the application of microwaves, gas flow, temperature monitoring(before, during, and after solvent evaporation), moderating microwaveenergy based upon the monitored temperature, and varying the microwaveenergy application time depending on the sample volume. The step ofapplying microwave energy to the sample while concurrently providing gasflow over and around the sample essentially dries the solvent from thechromatography media. The step of creating a gas flow further comprisescreating a vacuum in the vessel. The method illustrated in FIG. 4 may beperformed manually or preferably fully automated.

The method of preparing samples for column chromatography includes thefield of chromatography known as flash chromatography. Flashchromatography is a simple, rapid form of preparative column liquidchromatography. Flash chromatography is useful for the rapid componentseparation of reaction products, for example, to test the percentcompletion of a given reaction under certain conditions, or to measurethe amount of reactants converted to products under various experimentalparameters.

The method includes applying microwave energy to a sample compositioncontaining at least one solvent having reactants dissolved therein. Theapplication of microwave energy encourages a chemical reaction andgenerates desired products. Thereafter, the microwave energy is turnedoff and an absorbent media is added to the sample to absorb the solvent.In a preferred embodiment, the media is compatible with the liquidchromatography that will subsequently separate the expected products.That is, the media is compatible with the solid and mobile phases usedin the flash chromatography step to separate, identify, and purify theproduct components. The absorbent media is selected from the groupconsisting of silica, sand, alumina, and diatomaceous earth.

The absorbent media is mixed with the sample in an amount sufficient toprovide a substantially dry mixture of the media and the sample, butless than an amount that overly broadens the resolution of the sampleduring liquid chromatography. For example, an insufficient amount ofchromatography media added to the sample and exposed to microwave energywill result in a viscous mixture from which the reaction products cannotbe easily separated. This tends to render the sample useless for furtheranalysis. An excess amount of chromatography media added to the sampleand exposed to microwave energy will negatively affect the resolution ofthe products in subsequent analysis. In a preferred embodiment, thevolume of absorbent media used is about equal to the volume of solventwith products dissolved therein.

In this embodiment, the chromatography media can be added to the vesselcontaining the sample, thus avoiding the need to transfer the samplebetween vessels.

Thereafter, the microwave energy is applied to the dry mixture of themedia and the sample to encourage the solvent to evaporate under theinfluence of the microwave energy. Next, the method includes adding thedry mixture of the media and the remaining sample to a liquidchromatography column and separating the components of the remainingsample for identification and purification purposes.

In this manner, the sample preparation method carries out the microwaveassisted reaction in a microwave transparent vessel, mixes absorbentmedia with the sample in the same vessel to produce the dry mixture inthe vessel, and applies microwave energy to the dry mixture to removethe solvent while the dry mixture remains in the same vessel.

In a preferred embodiment, the method is automated and controlled by amicroprocessor. Furthermore, with the use of microwave energy for bothproduct synthesis and evaporation of solvent from the chromatographymedia, the microprocessor controls a cooling fan to actively cool thesample and prevent overheating of internal components.

In another aspect, the invention is a method of preparing samples forcolumn chromatography, including flash chromatography, and includesadding a scavenging composition to the sample to remove contaminants andexcess reagents.

Scavenging compositions include, but are not limited to, electrophilescavengers, nucleophile scavengers, base scavengers, acid scavengers.

The method further includes adding a coupling reagent to the sample tofacilitate the synthesis of products and adding a catalyst reagent tothe sample to accelerate the rate of the reaction. These steps are leftto the discretion of the researcher.

Scavengers, coupling reagents, and catalysts are known to one ofordinary skill in the art of organic chemical synthesis. In chemistry, ascavenger is defined as any substance added to a system or mixture toconsume or inactivate traces of impurities (Hawley's Condensed ChemicalDictionary, Twelfth Edition (1993); Van Nostrand Reinhold). Extensive,but not limiting, lists of scavengers, coupling reagents, catalysts, andother reagents acceptable for use with the method of the invention arefound in the Stratospheres™ Synthesis and Purification Guide publishedby Polymer Laboratories and the Organic Synthesis and PurificationCatalog (2003-2004) published by Silicycle™. Some reagents mentioned inthe catalogs are bound to a solid phase resin, such as silica, forconvenience. It should be known, however, that the reagents do not needto be bound to a solid phase to be chemically active.

The method of the invention further includes accelerating the steps ofscavenging, coupling, and catalyzing (collectively or individually) withmicrowave energy.

In yet another aspect, the invention is a method of dry-loading samplesfor column chromatography. The method includes adding an amount ofchromatography media to a vessel holding a sample, the sample beingdissolved in a solvent. The solvent is then evaporated using microwaveenergy, leaving dried chromatography media containing the sample. Thedried chromatography media containing the sample is thereafterdry-loaded onto a separation chromatography column (such as a flashchromatography column) for identification and purification of the samplecomponents.

The steps of adding chromatography media to the vessel, evaporating thesolvent using microwave energy, and loading the dried chromatographymedia containing the sample onto a separation chromatography column areperformed manually, or in a preferred embodiment, performedautomatically.

For example, and with respect to the instrument 10 of the invention,FIG. 5 depicts the instrument 10 with the NAVIGATOR™ peripheral device59 attached. The NAVIGATOR™ device 59 in combination with the instrument10 of the present invention provides one embodiment of the method forautomatic microwave assisted organic chemical synthesis andpurification. The automated aspect of the NAVIGATOR™ 59 is typicallyaided by a robotic arm 60.

By way of example, the NAVIGATOR™ device 59 in combination with theinstrument 10 of the present invention will begin chemical synthesis byplacing liquid reactants dissolved in a reaction solvent from the liquidhandling pumps 61 into the microwave transparent vessel 20. Couplingreagents, catalysts, and liquid scavenging compositions may also beadded at this time. Next, the vessel top portion (not shown) is placedonto the vessel 20 at the capping/decapping station 62. The robotic arm60 then moves the vessel 20 to the microwave cavity 19 of the instrument10 where microwave energy encourages the conversion of reactants toproducts. Following product formation, the robotic arm 60 moves thevessel 20 to the capping/decapping station 62, and removes the vesseltop portion. If necessary, a scavenging agent may be added at this time.If the scavenging agent is a liquid, the vessel 20 is placed under aliquid handling pump 61. If the scavenging composition is a solid (e.g.,a scavenging molecule attached to a solid phase media), the vessel 20 ismoved to the chromatography media reservoir 63. In a preferredembodiment, the scavenging composition is a solid as previouslydescribed and the media is an absorbent chromatography media selectedfrom the group consisting of silica, sand, alumina, and diatomaceousearth. The vessel top portion is then replaced.

The step of scavenging to remove contaminants and excess reagents isaccelerated by the application of microwave energy. Thus, the vessel 20is moved back to the microwave cavity 19 for exposure to microwaveenergy. In some embodiments, the reaction solvent is concurrentlyevaporated, while in other embodiments microwave energy is applied tothe scavenger-sample mixture before the sample is mixed with thechromatography media. Solvent evaporation is aided using one of the twopreviously discussed mechanisms for moving gas through the vessel, e.g.,the gas pump (not shown) will maintain a vacuum in the vessel tofacilitate the evacuation of solvent, or the gas pump or a gas reservoir(not shown) will maintain pressure in the vessel to facilitate theevacuation of solvent. Solvent evaporation leaves dried chromatographymedia containing the sample.

Following evaporation of the solvent from the dry mixture, the driedchromatography media is dry-loaded onto a separation chromatographycolumn (not shown), such as a flash chromatography column, and theproduct components are separated for identification and purificationpurposes.

The method appreciates the same benefits of microprocessor control aspreviously described, such as automatic control of at least the steps ofcarrying out the reaction, applying microwave energy to the sample, .The microprocessor may further “pulse” the microwave energy aspreviously described.

FIG. 6 is a rear perspective view of the instrument housing 11 thatillustrates some additional items. FIG. 6 illustrates the cooling fan43, a power switch 45, and a power cord inlet 47. In order to takeadvantage of the full capacity of the instrument, in preferredembodiments, the instrument includes a parallel port 49 and a serialport 50 for receiving input from or providing output to other electronicdevices, particularly microprocessor based devices, such as the pump 46(FIG. 3), personal computers, personal digital assistants or otherappropriate devices (not shown). Similarly, FIG. 6 illustrates aconnector 51 for the pressure transducer 37 previously described.

In the specification and the drawings, typical and preferred embodimentsof the invention have been disclosed. Specific terms have been used onlyin a generic and descriptive sense, and not for purposes of limitation.The scope of the invention is set forth in the following claims.

1. A method of preparing samples for column chromatography comprising:applying microwave energy to a sample composition containing at leastone solvent to encourage a chemical reaction and generate desiredproducts; thereafter mixing an absorbent media with the sample to absorbthe solvent, the media being compatible with liquid chromatography thatwill separate the expected products, being chemically inert to theexpected products, and being added in an amount sufficient to provide asubstantially dry mixture of the media and the sample, but less than anamount that overly broadens the resolution of the sample during liquidchromatography; thereafter applying microwave energy to the dry mixtureof the media and the sample to thereby encourage the solvent toevaporate under the influence of the microwave energy; and thereafteradding the dry mixture of the media and the remaining sample to a liquidchromatography column and separating the components of the remainingsample for identification and purification purposes.
 2. A samplepreparation method according to claim 1 comprising: carrying out themicrowave assisted reaction in a microwave transparent vessel; mixingthe absorbent media with the sample in the same vessel to produce thedry mixture in the vessel; and applying the microwave energy to the drymixture to remove the solvent while the dry mixture remains in the samevessel.
 3. The method as in claim 2 comprising computer microprocessorcontrol of at least the steps of carrying out the reaction and applyingmicrowave energy to the sample.
 4. The method as in claim 3 wherein thestep of carrying out a reaction comprises using pulsed microwave energycontrolled by the microprocessor.
 5. The method as in claim 1 whereinthe sample is actively cooled using a cooling fan.
 6. The method as inclaim 1 wherein the absorbent media is selected from the groupconsisting of silica, sand, alumina, and diatomaceous earth.
 7. Themethod as in claim 6 wherein the absorbent media comprises a volume ofmedia about equal to the volume of solvent.
 8. A method of preparingsamples for column chromatography, comprising: applying microwave energyto a sample composition containing at least one solvent to encourage achemical reaction and generate desired products; mixing a scavengingcomposition with the sample to remove contaminants and excess reagents;mixing an absorbent media with the sample to absorb the solvent, themedia being compatible with liquid chromatography that will separate theexpected products, being chemically inert to the expected products, andbeing added in an amount sufficient to provide a substantially drymixture of the media and the sample, but less than an amount that overlybroadens the resolution of the sample during liquid chromatography;thereafter applying microwave energy to the dry mixture of the media andthe sample to thereby encourage the solvent to evaporate under theinfluence of the microwave energy; and thereafter adding the dry mixtureof the media and the remaining sample to a liquid chromatography columnand separating the components of the remaining sample for identificationand purification purposes.
 9. The method of claim 8 wherein the step ofadding a scavenging composition to the sample comprises electrophilescavenging compositions, nucleophile scavenging compositions, basescavenging compositions, and acid scavenging compositions.
 10. Themethod of claim 8 further comprising the step of applying microwaveenergy to the scavenger-sample mixture to accelerate the scavengingremoval of contaminants and excess reagents prior to the step of mixingthe absorbent media with the sample.
 11. The method of claim 8comprising adding a coupling reagent to the sample to facilitate theconversion of reactants to products.
 12. The method of claim 8comprising adding a catalyst reagent to the sample to accelerate therate of the reaction.
 13. The method of claim 8 wherein columnchromatography comprises the technique of flash chromatography.
 14. Amethod of dry-loading samples for column chromatography, comprising:adding an amount of chromatography media to a vessel holding a sample,the sample being dissolved in a solvent; evaporating the solvent usingmicrowave energy, leaving dried chromatography media containing thesample; and dry-loading the dried chromatography media containing thesample onto a separation chromatography column for identification andpurification of the sample components.
 15. The method according to claim14 wherein the step of adding an amount of chromatography media to avessel comprises an amount about equal to the amount of solvent.
 16. Themethod according to claim 14 wherein the step of adding an amount ofchromatography media to the vessel comprises manually adding thechromatography media to the vessel.
 17. The method according to claim 14wherein the step of loading the dried chromatography media containingthe sample onto a separation chromatography column comprises manuallyloading the chromatography media onto the column.
 18. The methodaccording to claim 14 wherein the step of adding an amount ofchromatography media to the vessel comprises a microprocessor inelectronic control of adding an amount of chromatography media to thevessel.
 19. The method according to claim 14 wherein the step ofevaporating the solvent using microwave energy comprises amicroprocessor in electronic control of the microwave energy.
 20. Themethod according to claim 19 wherein the step of evaporating the solventusing microwave energy comprises a microprocessor in electronic controlof pulsing the microwave energy.
 21. The method according to claim 14wherein the step of dry-loading the dried chromatography mediacontaining the sample onto a separation chromatography column comprisesa computer microprocessor in electronic control of loading the driedchromatography media containing the sample onto a separationchromatography column for identification and purification of the samplecomponents.